MOLECULAR-CLONING OF A MOUSE MELANOCORTIN-5 RECEPTOR GENE WIDELY EXPRESSED IN PERIPHERAL-TISSUES

A mouse genomic clone named HGMP01B has been isolated by homology scre ening with a probe representing part of the human melanocortin 3 recep tor gene. HGMP01B was found to encode a 325 amino acid protein with al l the landmarks of G-protein-coupled receptors and belonging to the gr owing melanocortin receptor family. This receptor displays four potent ial sites for N-linked glycosylation and five potential sites of phosp horylation by protein kinase C. The HGMP01B gene was found to be expre ssed in many tissues, including skin, adrenal gland, skeletal muscle, bone marrow, spleen, thymus, gonads, uterus, and brain. A stable Chine se hamster ovary (CHO) cell line expressing approximately 10 000 recep tors per cell was established. This cell line displayed a saturable bi nding capacity for the radioiodinated alpha-melanocyte-stimulating hor mone (alpha-MSH) analog Nle(4),D-Phe(7)-alpha-MSH (NDP-MSH) with an apparent K-d of 1.47 +/- 0.15 nM. Binding of the labeled ligand was co mpeted for by all melanocortin peptides, except beta-endorphin or cort icotropin-like intermediate lobe peptide (CLIP). NDP-MSH was the most powerful competitor, followed by alpha-MSH, adrenocorticotropic hormon e (ACTH), beta-MSH, the gamma-MSHs, and ACTH 4-10. Functional assays c onfirmed that HGMP01B, like other melanocortin receptors, stimulated a denylyl cyclase. The potency order obtained in these cyclic adenosine monophosphate (cAMP) accumulation assays was consistent with that of t he binding studies. HGMP01B therefore appears as a fifth melanocortin receptor (MC5), responding mainly to alpha-MSH (EC(50) = 1.07 +/- 0.13 nM) and endowed with a pharmacological profile similar to that of the melanocyte MSH (MC1) receptor, but characterized by a broad tissue di stribution. The expression of MC5 in lymphoid organs suggests that thi s receptor could be implicated in the reported antiinflammatory action of melanocortins.